The emergence of severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome (MERS)-CoV underscores the threat of cross-species transmission events leading to outbreaks in humans. Here we examine the disease potential of a SARS-like virus, SHC014-CoV, which is currently circulating in Chinese horseshoe bat populations1. Using the SARS-CoV reverse genetics system2, we generated and characterized a chimeric virus expressing the spike of bat coronavirus SHC014 in a mouse-adapted SARS-CoV backbone. The results indicate that group 2b viruses encoding the SHC014 spike in a wild-type backbone can efficiently use multiple orthologs of the SARS receptor human angiotensin converting enzyme II (ACE2), replicate efficiently in primary human airway cells and achieve in vitro titers equivalent to epidemic strains of SARS-CoV. Additionally, in vivo experiments demonstrate replication of the chimeric virus in mouse lung with notable pathogenesis. Evaluation of available SARS-based immune-therapeutic and prophylactic modalities revealed poor efficacy; both monoclonal antibody and vaccine approaches failed to neutralize and protect from infection with CoVs using the novel spike protein. On the basis of these findings, we synthetically re-derived an infectious full-length SHC014 recombinant virus and demonstrate robust viral replication both in vitro and in vivo. Our work suggests a potential risk of SARS-CoV re-emergence from viruses currently circulating in bat populations.

Main

The emergence of SARS-CoV heralded a new era in the cross-species transmission of severe respiratory illness with globalization leading to rapid spread around the world and massive economic impact3,4. Since then, several strains—including influenza A strains H5N1, H1N1 and H7N9 and MERS-CoV—have emerged from animal populations, causing considerable disease, mortality and economic hardship for the afflicted regions5. Although public health measures were able to stop the SARS-CoV outbreak4, recent metagenomics studies have identified sequences of closely related SARS-like viruses circulating in Chinese bat populations that may pose a future threat1,6. However, sequence data alone provides minimal insights to identify and prepare for future prepandemic viruses. Therefore, to examine the emergence potential (that is, the potential to infect humans) of circulating bat CoVs, we built a chimeric virus encoding a novel, zoonotic CoV spike protein—from the RsSHC014-CoV sequence that was isolated from Chinese horseshoe bats1—in the context of the SARS-CoV mouse-adapted backbone. The hybrid virus allowed us to evaluate the ability of the novel spike protein to cause disease independently of other necessary adaptive mutations in its natural backbone. Using this approach, we characterized CoV infection mediated by the SHC014 spike protein in primary human airway cells and in vivo, and tested the efficacy of available immune therapeutics against SHC014-CoV. Together, the strategy translates metagenomics data to help predict and prepare for future emergent viruses.

抽象的

严重急性呼吸综合征冠状病毒 (SARS-CoV) 和中东呼吸综合征 (MERS)-CoV 的出现凸显了导致人类爆发的跨物种传播事件的威胁。在这里，我们研究了一种 SARS 样病毒 SHC014-CoV 的疾病潜力，该病毒目前在中国马蹄蝠种群1 中传播。使用 SARS-CoV 反向遗传学系统2，我们生成并表征了一种嵌合病毒，该病毒在小鼠适应的 SARS-CoV 骨架中表达蝙蝠冠状病毒 SHC014 的尖峰。结果表明，在野生型骨架中编码 SHC014 尖峰的 2b 组病毒可以有效地使用 SARS 受体人血管紧张素转化酶 II (ACE2) 的多个直向同源物，在原代人气道细胞中有效复制，并实现与流行病相当的体外滴度SARS-CoV 病毒株。此外，体内实验证明嵌合病毒在小鼠肺中的复制具有显着的发病机制。对现有的基于 SARS 的免疫治疗和预防方法的评估显示疗效不佳；使用新型刺突蛋白，单克隆抗体和疫苗方法均未能中和并保护其免受 CoV 感染。在这些发现的基础上，我们重新合成了一种传染性全长 SHC014 重组病毒，并在体外和体内证明了强大的病毒复制。我们的工作表明，目前在蝙蝠种群中传播的病毒存在重新出现 SARS-CoV 的潜在风险。

主要的

SARS-CoV 的出现预示着严重呼吸道疾病跨物种传播的新时代，全球化导致了全球范围内的快速传播和巨大的经济影响3、4。从那时起，动物种群中出现了几种毒株，包括甲型流感毒株 H5N1、H1N1 和 H7N9 以及 MERS-CoV，给受灾地区造成了相当大的疾病、死亡率和经济困难5。尽管公共卫生措施能够阻止 SARS-CoV 的爆发4，但最近的宏基因组学研究已经确定了在中国蝙蝠种群中传播的密切相关的 SARS 样病毒序列，这些病毒可能对未来构成威胁1 , 6. 然而，仅凭序列数据就无法识别和准备未来的大流行前病毒。因此，为了检查循环蝙蝠 CoV 的出现潜力（即感染人类的潜力），我们构建了一种嵌合病毒，该病毒编码一种新型的人畜共患 CoV 刺突蛋白——来自从中国马蹄蝠中分离的 RsSHC014-CoV 序列1 ——在 SARS-CoV 小鼠适应骨干的背景下。杂交病毒使我们能够独立于其天然骨架中其他必要的适应性突变来评估新型刺突蛋白引起疾病的能力。使用这种方法，我们在原代人类呼吸道细胞和体内表征了由 SHC014 刺突蛋白介导的 CoV 感染，并测试了现有免疫疗法对 SHC014-CoV 的功效。该策略共同转化宏基因组学数据，以帮助预测和准备未来出现的病毒。